RESUMEN
AIM: To determine whether chemokines such as SDF-1 and monocyte chemoattractant protein-1 (MCP-1) are responsible for hydrogen peroxide (H2 O2 )-induced extracellular matrix (ECM) degradation and to identify the underlying mechanism in human dental pulp cells (HDPCs). METHOD: Human dental pulp cells were exposed to 0.4 mmol H2 O2 for 48 h. mRNA expression and protein expression were examined by RT-PCR and Western blot analysis, respectively. The mRNA expression of chemokine (SDF-1 and MCP-1), their receptors (CXCR4 and CXCR2) and extracellular matrix proteins was evaluated by reverse transcriptase-polymerase chain reaction. The production of SDF-1, MCP-1, CXCR4 and CCR2 in the culture medium was determined by enzyme-linked immunosorbent assay. Signal transduction pathway was examined by Western blotting. RESULTS: Hydrogen peroxide provoked the activation of MCP-1 and SDF-1 mRNA and their respective receptors, CXCR4 and CXCR2. H2 O2 treatment concomitantly downregulated the expression of ECM molecules, such as type I collagen, elastin and fibronectin, and upregulated the mRNA expression of matrix metalloproteinase-1 (MMP-1), MMP-2, MMP-8 and MMP-9. Hydrogen peroxide-induced ECM degradation and MMP upregulation were blocked by neutralizing antibodies and siRNAs directed against SDF-1 and MCP-1. Inhibition of SDF-1 and MCP-1 blocked the H2 O2 -induced activation of Akt, p38, ERK and NF-kB. CONCLUSION: Inhibition of SDF and MCP-1 is a potent component of reducing release reactive oxygen species-induced ECM degradation in HDPCs and may play an important role in pulpal and periapical inflammation.
Asunto(s)
Quimiocina CCL2/metabolismo , Quimiocina CXCL12/metabolismo , Pulpa Dental/citología , Matriz Extracelular/efectos de los fármacos , Matriz Extracelular/metabolismo , Peróxido de Hidrógeno/farmacología , Western Blotting , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Humanos , Metaloproteinasas de la Matriz/metabolismo , ARN Mensajero/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de SeñalRESUMEN
Gemin5 is a 170-kDa WD-repeat-containing protein that was initially identified as a component of the survival of motor neurons (SMN) complex. We now show that Gemin5 facilitates the activation of apoptosis signal-regulating kinase 1 (ASK1) and downstream signaling. Gemin5 physically interacted with ASK1 as well as with the downstream kinases SEK1 and c-Jun NH(2)-terminal kinase (JNK1), and it potentiated the H(2)O(2)-induced activation of each of these kinases in intact cells. Moreover, Gemin5 promoted the binding of ASK1 to SEK1 and to JNK1, as well as the ASK1-induced activation of JNK1. In comparison, Gemin5 did not physically associate with MKK7, MKK3, MKK6, or p38. Furthermore, depletion of endogenous Gemin5 by RNA interference (RNAi) revealed that Gemin5 contributes to the activation of ASK1 and JNK1, and to apoptosis induced by H(2)O(2) and tumor necrosis factor-alpha (TNFalpha) in HeLa cells. Together, our results suggest that Gemin5 functions as a scaffold protein for the ASK1-JNK1 signaling module and thereby potentiates ASK1-mediated signaling events.
Asunto(s)
MAP Quinasa Quinasa Quinasa 5/metabolismo , Proteína Quinasa 8 Activada por Mitógenos/metabolismo , Ribonucleoproteínas Nucleares Pequeñas/metabolismo , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Secuencia de Bases , Línea Celular , ADN Complementario/genética , Células HeLa , Humanos , Peróxido de Hidrógeno/farmacología , Técnicas In Vitro , MAP Quinasa Quinasa Quinasa 5/genética , Proteína Quinasa 8 Activada por Mitógenos/genética , Unión Proteica , ARN Interferente Pequeño/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Ribonucleoproteínas Nucleares Pequeñas/antagonistas & inhibidores , Ribonucleoproteínas Nucleares Pequeñas/genética , Proteínas del Complejo SMN , Transducción de Señal , Transfección , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
We used the IL088 Otodynamic Analyzer system to study click-evoked otoacoustic emissions (CEOAEs) in 30 healthy guinea pigs. The animals were anesthetized and patterns of the CEOAEs were evaluated before manipution, after the tympanic bulla was opened, and after formation of a microfistula on the basal turn of the cochlea. The animals then were divided into three pressure loading groups (10, 20, and 30 cm H2O). CEOAEs were recorded with a capillary manometer at pretest, 5, 10, 20, 30, 45, and 60 min after perilymphatic pressure loading to the basal turn of cochlea, and 10 and 20 min after pressure unloading. As perilymphatic pressure increased, all three pressure groups showed maximum decreases in both echo response and reproducibility 5 min after pressure loading. In the 10 cm H2O pressure group, emissions recovered 10 min after pressure loading, and this tendency continued. However, in the 20 and 30 cm H2O pressure groups, no recovery of emissions was seen throughout the 60 min observation period, except for emissions after pressure unloading. The results suggest that the echo response and reproducibility may be sensitive indicators of cochlear function and perilymphatic pressure regulation capacity.
Asunto(s)
Estimulación Acústica , Cóclea/fisiología , Cobayas , Perilinfa/fisiología , AnimalesRESUMEN
From a nationwide survey of otitis media in Korea, 44.52% of the population were found to have some type of otitis media or its sequelae. A high prevalence rate was seen in the age group over 41 years. This finding suggests a strong relationship between socioeconomic status and incidence of otitis media. From a clinical study of surgical cases of otitis media seen in the past 10 years, we have found that the prevalence of chronic otitis media is decreasing every year. However, severity and pathological findings of otitis media were reflected remarkably in a decreased incidence of acute purulent otitis media and an increased incidence of middle ear effusion in children. In recent years our efforts to control chronic otitis media in children have focused on the treatment of chronic middle ear effusion. To prevent the latter condition, it is strongly emphasized that pediatricians and primary care physicians should be competent in diagnosing otitis media as early as possible, and that they should refer appropriate patients to otolaryngologists for further evaluation and management.